Abstract
Fresh fruits and vegetables promote good health but harbour a wide range of microbial contaminants. To assess the microbial quality of fruits and vegetables, twenty-eight (28) samples of different fruits and vegetables were purchased from various vendors. Samples were analyzed to study the density of microorganisms. The spread plate method was used for the isolation of bacteria on nutrient media. Mean bacterial load ranged from 4.4x106cfu/g to 1.07x107cfu/g with garden eggs recording the lowest count and cabbage samples harbouring more bacteria than the rest of the samples. Sliced watermelon samples recorded a mean viable count of 9.2x106cfu/g. Mean coliform count was highest in cabbage (7.4x104cfu/g) and lowest in pineapple (1.1x104cfu/g). Six bacteria belonging to eight genera were identified. Staphylococcus aureus (82.1%) was the most frequently isolated followed by Salmonella species 13(46.4%), Bacillus species 12(42.9%), Enterobacter aerogenes 9(32.1%). Micrococcus species 6(21.4%) and Escherichia coli (25%) were the least frequently isolated. Resistance to a wide range of antibiotics was observed. Resistance to augmentin, cefuroxime, ceftazidime and ceftriaxone was observed as 100%, 100%, 100% and 69.2% respectively for the Salmonella isolates. Drug susceptibility assay revealed that 84.6%, 69.2%, 61.5% and 30.8% of the Salmonella isolates investigated were susceptible to ciprofloxacin, Ofloxacin, gentamicin and ceftriaxone respectively. Eighty-seven percent of the S. aureus isolates were resistant against erythromycin whereas 82.6% and 73.9% of the isolates were resistant to augmentin and ceftriaxone respectively. Resistance to ofloxacin was seen in 14.3% while 100% were resistant to Augmentin for the E. coli isolates. The presence of potential food-borne pathogens makes the fruits to be microbiologically unsafe and they should be pre- treated thoroughly before human consumption, so as to reduce the risk of food- borne outbreaks.
TABLE OF CONTENT
Cover
page i
Title
page ii
Certification
iii
Declaration
iv
Dedication
v
Acknowledgments vi
Table
of contents vii
List
of Tables x
Abstracts xi
CHAPTER ONE
1.1 Introduction 1
1.2 Aims and Objectives 6
CHAPTER TWO: LITERATURE REVIEW
2.1 An
Overview of Fruits 7
2.2 Bacteria in Fruits as a
cause of Disease 8
2.2.1 Risk associated with consumption of fruits
(Microbial hazards) 9
2.3 Microbial Contamination 11
2.3.1 Soft Rot 12
2.4 Contamination Sources of Vegetables 12
2.4.1 Pre-Harvest Contamination 13
2.4.2 Quality of Irrigation Water 14
2.4.3 Post-Harvest Contamination 15
2.5 Common
Fruits and Vegetable Spoilage Organisms 16
2.5.1 Listeria Monocytogenes 16
2.6 Global Relevance of Food
Safety and Foodborne Illness 17
2.7 Food
Poisoning 20
2.8 Antibiotics
Resistance 21
CHAPTER
THREE: MATERIAL AND METHODS
3.1 Sample Collection 23
3.2 Media
Preparation 23
3.3 Preparation of Sample Homogenate 23
3.3.1 For
The Isolation Of Salmonella Species 24
3.4 Gram Staining 25
3.5 Biochemical Tests 25
3.5.1 Catalase Test 25
3.5.2 Methyl Red
Test 26
3.5.3 Voges- Proskauer Test 26
3.5.4 Indole Test 26
3.5.5 Citrate
Utilization Test 27
3.5.6 Hydrogen Sulphide (H2s) Production
Test 27
3.6 Antibiotics
Sensitivity Test 27
CHAPTER FOUR
Results 29
CHAPTER
FIVE
5.1 Discussion 35
5.2 Conclusion 40
REFERENCES 41
LIST
OF TABLES
Table 4.1: Mean
Bacterial and Fungal Load in cfu/g of the Samples 30
Table 4.3 Frequency
of Isolation of the Microorganisms 31
Table 4.2: Colonial Morphology and Biochemical Tests for
Identification of the Bacterial Isolates 32
Table 4.5: Distribution
of the Isolates across the various Samples 33
Table 4.4: Antibiotic
Susceptibility Profile of the Isolates 34
CHAPTER ONE
1.1 INTRODUCTION
The sales of fresh-cut fruit and vegetables have considerably
increased in Nigeria over the last two decades (Gorny, 2005; James and
Ngarmsak, 2010). The increase in consumption of fresh-cut fruit is mostly
because people are more aware of the importance and benefits of healthy eating
habits and have less time for food preparation. Fresh-cut produce fit the many
needs of a modern lifestyle as they serve colourful, flavourful and nutritional
(energy, vitamins, minerals, and dietary fibre) compounds and are also convenient
to use and consume (Kader and Barrett, 2005).
However, fresh-cut fruit and vegetables are highly perishable
products and have a very short shelf-life, even at chill temperatures mainly
due to water loss, translucency, browning, softening, surface dehydration,
off-flavour and off-odour development, and microbial spoilage (Rojas-Grau et al., 2009). The fast deterioration of
fresh-cut fruits and vegetables results mostly from the damage caused to the
cells and tissues by cutting and trimming and the removal of their natural
protective barriers (Olivas et al.,
2005). Cutting and/or peeling may induce ripening, cause senescence and aid the
rapid growth of contaminating microorganisms due to the release of nutrients
from the damaged cells and tissues. Therefore, the spoilage phenomena on
fresh-cut fruit are related to biochemical processes, both in microorganisms
and in the fruit tissue itself.
Food safety is the assurance that food will not cause any
harm to the consumer when it is prepared and/or consumed according to its
intended use. Ready to eat (RTE) fruit constitute a suitable and convenient
meal for today’s lifestyles, because they need no cooking or further
preparation. As well as being considered low-calorie food, they are rich in
fiber and provide a great variety of vitamins, minerals, and other
phyto-chemicals (Sarjo et al., 2006). However fruits are widely exposed
to microbial contamination through contact with soil/dust and water and poor
handling at harvest or during postharvest processing. They, therefore create
favorable condition for diverse range of microorganisms including plant and
human pathogens (Nguyen and Carlin, 1994).
The causative agents of microbiological spoilage in fruits
can be bacteria, as well as yeasts and molds. The main spoilage agents can be
considered as due to the low pH of most fruits. Some bacteria such as
Campylobacter spp., E. coli O157:H7, Salmonella spp., Listeria
monocytogenes, Staphylococcus aureus, Shigella spp, Erwinia spp.,
Enterobacter spp., Alicyclobacillus spp., Propionibacterium
cyclohexanicum, Pseudomonas spp., and lactic acid bacteria can cause
spoilage in fruit (Walker and Phillips, 2008). Certain common molds such as Penicillium
spp., Aspergillus spp., Eurotium spp., Alternaria spp.,
Cladosporium spp., Paecilomyces spp., and Botrytis spp.
have been shown to be involved in the spoilage of fresh fruits (Lund and
Snowdon, 2000).
Most microorganisms that are initially observed on whole fruit
or vegetable surfaces are soil inhabitants, members of a very large and diverse
community of microbes that collectively are responsible for maintaining a
dynamic ecological balance within most agricultural systems. Vectors for
disseminating these microbes include soil particles, airborne spores, and
irrigation water. Most bacteria and fungi that arrive on the developing crop
plant either are completely benign to the crop’s health or, in many instances,
provide a natural biological barrier to infestation by the subset of
microorganisms responsible for crop damage (Andrews and Harris, 2000). Microbial
profile of fruits and vegetables are direct reflection of the sanitary quality
of the cultivation, harvesting, transportation, storage, and processing of the
produce (Janisiewicz and Korsten, 2002; Andrew and Harris, 2000). The
difference in the microbial profiles of fruits also result largely from
unrelated factors like resident micro-flora in the soil and nonresident
micro-flora through animal manures, sewage or irrigation water, transportation
and handling by sellers (Ray and Bhunia, 2007; Ofor et al., 2009).
Microbial infections of food borne origin
are a major public-health problem internationally and a significant cause of
death in developing countries (WHO, 2006). Food safety in developing countries
is influenced by a number of factors.
There is a high risk of contamination at
all stages of production, processing and distribution which are very difficult
to control through regulations given the common constraints in supporting
infrastructure and institutional capacities. Quantitative microbial risk
assessment can help in identifying critical control points (Seidu et al.,
2008). These microorganisms are carried on hands, wiping cloths and utensils,
especially chopping boards. The slightest contact can transfer them to food and
cause food borne disease.
The majority of microorganisms associated
with raw vegetables are non-pathogenic and gram negative organisms tend to
dominant the bacterial population including Enterobacter spp. and other
coliforms (Janisiewicz and Korsten, 2002).
Microbiological contamination of fruit can occur directly or indirectly from
animals or insects, soil, manures, water and equipment used to grow the
horticultural commodities as well as human handling along the food chain. The
microbiological contaminants may have an adverse health effect.
Fresh vegetables normally carry natural
non-pathogenic epiphytic microorganisms, however, during growth, harvest,
transportation, and further handling the produce can be contaminated with
pathogens from animal and human sources (Morgante et al., 2008). As most of these produce are eating without
further processing, their microbial content may represent a risk factor for the
consumer’s health (Ragaert et al.,
2007).
Consumption of fruit and vegetable
products is commonly viewed as a potential risk factor for infection with
enteropathogens such as Salmonella and Escherichia coli O157,
with recent outbreaks linked to lettuce, spinach and tomatoes.
More than 90 percent of the cases of food
poisoning each year are caused by Staphylococcus aureus, Salmonella,
Clostridium perfringens, Campylobacter, Listeria monocytogenes, Vibrio
parahaemolyticus, Bacillus cereus, and Enteropathogenic Escherichia coli
and Proteus. Fresh vegetables can be a vehicle for transmission of
bacterial, parasitic and viral pathogens capable of causing human illness and a
number of reports refer to raw vegetables, harboring potential food borne
pathogens Escherichia coli, Listeria monocytogenes, Sallmonella
was isolated from raw vegetables (Nweze, 2010).
It has been shown that Street-vended foods
have been implicated in outbreaks of foodborne illnesses all around the world
(Bryan et al., 1992). Most food-related illnesses have historically been
attributed to one of five major groups of pathogenic bacteria (Mboto et al., 2012). These five groups are Salmonella,
Shigella, Clostridium botulinum, Clostridium perfringens, Bacillus
cereus, and Staphylococcus aureus. These have been joined by the
emerging pathogens such as Yersinia enterocolitica, Escherichia coli,
Listeria monocytogens, and Campylobacter jejuni (Mboto et al., 2012).
Foodborne diseases of microbiological
origin can be caused by a variety of agents, which gain entry by the
gastrointestinal tract. However the symptoms are often mild and self-limiting.
Symptoms of foodborne disease, which are not necessarily confined to diarrhoea
and vomiting, are caused by viable organisms and/or by the toxins that they
produce. The risk of disease from these agents varies depending on the
pathogen, the dose, the host and the properties of the food matrix. Host risk
factors include age, immune status, underlying debilitating disease or stress
factors, and the physiological state of the stomach and upper small intestine
at the time of exposure to the agent. For these reasons a minimum infectious
dose cannot be defined, although the risk of disease at low exposure for some
agents is small (Nweze, 2010). The presence of foodborne agents that may cause
illness in ready-to-eat foods is a significant risk to consumer health and
their absence is of paramount importance.
With the exception of the aerobic and
anaerobic bacterial spores, detection of foodborne pathogenic agents at any
level is of concern and should be investigated with an urgency of response
proportionate to the level of contamination and risk to consumers. Although low
numbers of pathogens, such as coagulase-positive staphylococci, C.
perfringens, B. cereus, and L. monocytogenes, in ready-to-eat
products probably represent a very low risk to immunocompetent people, they are
more significant for the immunocompromised and vulnerable groups. Low levels
may be due to natural contamination of raw materials used in those foods, but
usually their presence suggests faults in the production or subsequent handling
of food which could lead to an unacceptable increase in risk. There may also be
a need for action when detecting low numbers of these organisms in ready-to-eat
foods because there is variation in host susceptibility and inter strain
differences in the pathogenicity of these bacteria.
In developing countries because of inadequate or even non
existing systems for routine diagnosis and monitoring or reporting for many of
the food-borne pathogens, most outbreaks caused by contaminated fruit and
vegetables go undetected and the incidence of their occurrence in food during
pre and postharvest is underestimated (Dorny et al., 2009). This is even
exacerbated by the increasing trend of antibiotics resistance. The excessive
use of over-the counter antibiotics has led to an increase in
antibiotic-resistant microbes in the environment (Silbergeld et al., 2008). These
antibiotic-resistant bacteria have made their way into the food chain, and
clinical treatment for infections caused by these bacteria has become crucial,
especially for immunocompromised patients (Silbergeld et al., 2008).
The recurrent episodes of food borne
illnesses with symptoms of gastro intestinal distress like diarrhea, vomiting,
abdominal cramp and nausea has remained a major cause of mortality and
morbidity in Nigeria.
1.2 Aims and Objectives
a. To
evaluate the total heterotrophic bacterial load on vegetable and fruits
surfaces.
b. To
isolate, identify and characterize the food-borne pathogens associated with vegetable and fruits
surfaces.
c. To
determine the antibiotics susceptibility profile of the isolates that will be
isolated from the vegetable and fruits surfaces.
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